Due to the process of X-inactivation, which occurs normally in diploid female cells, the mammalian X chromosome offers unique opportunities to investigate the relationship between chromatin structure and gene expression. Most of the inactive X chromosome assumes the properties of heterochromatin, but certain genes escape inactivation and thus remain expressed from both X chromosomes in female cells. The recent description of six genes that escape clustered in a domain on Xp11.21-Xp11.22 supports the hypothesis that genes that escape are flanked by boundary elements that prevent the X-inactivation signal from spreading through them. Therefore, the experiments described in this proposal have three specific aims: (1) to construct a detailed physical map of this multigene domain of less than 370 kb; (2) to produce a map of the scaffold/matrix attachment sites of the chromatin in and around this domain; (3) to assemble a map of the chromatin conformation in the region of the domain on both the active and inactive X chromosome. The data generated by these experiments will allow for the meaningful correlation of primary sequence and chromatin structure on a unique region of the X chromosome, and may allow for the identification of boundary elements that function in the process of X-inactivation.